Sources and Consequences of noise in cell cycle regulation

细胞周期调节中噪音的来源和后果

• 批准号: 7479185
• 负责人: FREDERICK R. CROSS
• 金额: $ 31.18万
• 依托单位: ROCKEFELLER UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-08-01 至 2010-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7479185
• 关键词: Address; Architecture; Behavior; Binding Sites; Biological Process; Cell Cycle; Cell Cycle Regulation; Cell Size; Cells; Complex; Computers; Coupling; Data; Data Set; Depth; Disadvantaged; Dissociation; Environment; Equation; Eukaryota; Eukaryotic Cell; Event; Gene Expression; Generations; Genes; Genetic; Genome; Image; Individual; Lead; Length; Light; Maps; Measurement; Measures; Methods; Microfluidics; Microscopy; Mitotic; Modeling; Molecular; Monitor; Noise; Nuclear; Numbers; Operative Surgical Procedures; Physiologic pulse; Ploidies; Population; Population Study; Proliferating; Property; Proteins; Pulse taking; Rate; Reagent; Regulation; Regulon; Reporter; Repression; Research Personnel; Resolution; Resources; Rest; Running; Saccharomyces cerevisiae; Saccharomycetales; Site; Source; Structure; Synthetic Genes; System; Technology; Testing; Time; Variant; Yeasts; analytical method; analytical tool; base; cell growth; cellular imaging; design; genetic pedigree; improved; insight; mathematical model; mutant; novel; programs; promoter; response; single cell analysis; size; technology development

DESCRIPTION (provided by applicant): Single-cell variation (noise) in the operation of genetic networks is an inevitable consequence of the small size of cellular systems, leading to small numbers of molecules (genes, mRNAs, proteins) that may cause fluctuations in the function of important genetic control circuits. Where such noise is deleterious, it is likely that noise suppression mechanisms have evolved; noise could also be beneficial in some cases, by allowing breakout from dynamical dead-ends or by diversifying a population to meet a changing environment. Despite the significance of this topic, empirical results on noise have so far been largely restricted to simple synthetic gene circuits. We propose analysis of single-cell variation in a much more complex and physiologically relevant situation: the robustness of cell cycle traversal and duration of cell cycle intervals in the eukaryote S. cerevisiae. We have developed a quantitative fully automated time- lapse fluorescent microscopy setup allowing semi-automated analysis of cell-cycle-regulated gene expression and relocation of cell cycle regulators, throughout the generation of complete multi-cell cycle pedigrees. This capability can be integrated with the enormous amount of information available about the regulation of the budding yeast cell cycle, the availability of genetic reagents to systematically perturb the cell cycle, and useful deterministic mathematical models of the cell cycle engine. We will test the hypothesis that variability in G1 length is due to bistability at cell cycle Start. We will generate quantitative single-cell data on the relationship of cell and nuclear size to cell cycle transition times, in wild-type and mutants, to provide constraints for models on nucleo-cytoplasmic size coupling. We will attack the question of what promoter architectures may lead to different levels of single-cell variability in gene expression, using a synthetic promoter strategy combined with single-cell imaging. We have obtained direct evidence for stochastic molecular variation in cell cycle Start; we will pursue this finding genetically and with stochastic modeling. To pursue these ideas in more dynamic depth, we have developed microfluidic methods that allow periodic pulses of gene expression in individual monitored cells as they proliferate from single cells into microcolonies. With this technology we will test the response of the cell cycle engine to brief forced expression of regulators, and looking for mode-locking and other informative dynamic behaviors. We will extend these analytical tools to mitotic regulation, testing the hypothesis that the complex design of the mitotic oscillator functions to suppress noise. The results should shed light on fundamental questions of robustness and dynamics of the cell cycle engine at single-cell resolution, and on the advantages and disadvantages of variability in the operation of a fundamental complex genetic circuit.

描述（由申请人提供）： 遗传网络运行中的单细胞变异（噪声）是细胞系统尺寸较小的必然结果，导致少量的分子（基因，mRNA，蛋白质）可能导致重要遗传控制电路的功能波动。在这种噪声有害的地方，很可能会发展噪声机制。在某些情况下，噪声也可能是有益的，它可以通过动态的死胡同或使人群多样化以满足不断变化的环境来有益。尽管该主题具有重要意义，但迄今为止，对噪声的经验结果很大程度上仅限于简单的合成基因回路。我们提出在更复杂和生理相关的情况下对单细胞变异的分析：细胞周期遍历和酿酒酵母中细胞周期间隔的持续时间的鲁棒性。我们已经开发了一种定量的全自动时间流失荧光显微镜设置，可以在整个完整的多细胞周期谱系中对细胞周期调节的基因表达和细胞周期调节剂的迁移的半自动分析。该能力可以与有关发芽酵母细胞周期调节的大量信息，遗传试剂对系统扰动细胞周期的可用性以及细胞周期发动机的有用的确定性数学模型的可用性。我们将检验以下假设：G1长度的变异性是由于细胞周期开始时的双疗法。我们将在野生型和突变体中生成有关细胞和核大小与细胞周期过渡时间之间关系的定量单细胞数据，以提供对核质质大小耦合的模型的约束。我们将使用合成启动子策略与单细胞成像相结合，攻击哪些启动子体系结构可能导致基因表达中不同水平的单细胞变异性。我们已经获得了细胞周期开始中随机分子变异的直接证据。我们将通过遗传和随机建模来追求这一发现。为了在更动态的深度中追求这些思想，我们开发了微流体方法，这些方法允许单个受监测细胞中的基因表达的周期性脉冲从单个细胞增殖到微菌落中。通过这项技术，我们将测试细胞周期引擎对调节器的强迫表达的响应，并寻找模式锁定和其他内容丰富的动态行为。我们将将这些分析工具扩展到有丝分裂调节，并检验以下假设：有丝分裂振荡器的复杂设计功能抑制噪声。结果应阐明单细胞分辨率下的细胞周期发动机的鲁棒性和动力学的基本问题，以及基本复杂遗传回路运行中可变性的优势和缺点。